Immunofluorescent analysis of p21 was performed on 70% confluent log phase U-2 OS cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0. 25% Triton™ X-100 for 10 minutes, and blocked with 5% BSA for 1 hour at room temperature. The cells were labeled with ABfinity™ p21 recombinant rabbit monoclonal antibody (Product # 701151) at a dilution of 1:500 in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Flour® 488 goat anti-rabbit IgG secondary antibody (Product # A11008) at a dilution of 1:400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 594 phalloidin (Product # A12381). Panel d is a merged image showing nuclear localization. The images were captured using a Nikon microscope at 20X magnification.
|Tested species reactivity||Human|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Peptide corresponding to amino acids 217–227 of human p21 protein|
|Contains||0.09% sodium azide|
|Storage Conditions||Maintain refrigerated at 2-8°C for up to 1 month. For long term storage store at -20°C|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1:100-1:500|
|Western Blot (WB)||1:500-1:5000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
This antibody is predicted to react with equine, mouse, rabbit and rat based on sequence homology.
ABfinity™ recombinant antibodies are rabbit monoclonal antibodies, unmatched for producing superior results. ABfinity™ antibodies are developed by immunizing animals, screening for functionality, cloning the immunogen-specific antibody genes into high-level mammalian expression vectors, produced on a large scale, and purified with Protein A.
ABfinity™ monoclonal antibodies resemble rabbit monoclonals isolated from serum or produced by hybridomas, but demonstrate greater specificity and sensitivity. Because ABfinity™ recombinant antibodies are derived from cloned DNA sequences of the heavy and light antibody chains, they are not susceptible to cell-line drift or lot-to-lot variation, thus allowing for peak specificity and performance.
Intact IgG appears on a non-reducing gel as ~150 kDa band and upon reduction generating a ~25 kDa light chain band and a ~50 kDa heavy chain.
Protein p21 is a tumor suppressor protein which is an inhibitor of Cyclin-dependent kinases (CDK) and is transcriptionally activated by p53. It is important in the response of cells to genotoxic stress and a major transcriptional target of p53 protein. The occurrence of p21 in nucleus executes binding and inhibiting the activity of cyclin dependent kinases Cdk1 and Cdk2, and blocks the transition from G1 phase into S phase or from G2 phase into mitosis after DNA damage. This enables the repair of damaged DNA. In the cytoplasm, p21 protein has an anti-apoptotic effect. It is able to bind to and inhibit caspase 3, as well as the apoptotic kinases ASK1 and JNK. P21 also exhibits a dual function in cancerogenesis. It acts as a tumor suppressor, prevents apoptosis, and acts as an oncogene
IP-MS enrichment of P27 KIP1 (LFQ intensity): P27 KIP1 was enriched 236-fold from HCT116 lysate compared to background proteins, using the optimized IP-MS workflow with Pierce MS-Compatible Magnetic IP Kit protein A/G (Part No. 90409) and P27 KIP1 antibody (Part No. 701151). The STRING database (www.string-db.org) was used to identify the protein interactor list. See more information on IP-MS verification of antibody selectivity. IP-MS validation info.
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